The present invention relates to the microbial production of (6R)-2,2,6-trimethylcyclohexane-1,4-dione (hereinafter referred to as levodione), a useful intermediate in the production of carotenoids, such as (3R,3xe2x80x2R)-zeaxanthin. More particularly, the present invention relates to a process for producing levodione of high purity and in high yield by the catalytic reaction of 2,6,6-trimethyl-2-cyclohlexene-1,4-dione (hereinafter referred to as ketoisophorone) with a specific yeast.
Levodione has previously been prepared through the reduction of the carbon-carbon double bond in ketoisophorone by contacting the ketoisophorone with baker""s yeast, such as Saccharomyces cervisiae, which functions as an enantioselective biocatalyst (Biotechnology of Vitamins, Pigments and Growth Factors, Ed. Erick J. Vandamme, page 71, Elsevier Applied Science, London and New York). However, baker""s yeast is not suitable for use in the industrial production of levodione because the yields are too low. In addition, the use of baker""s yeast in the production of levodione is inefficient because the yeast cells cannot be reused because of the short lifetime of the reaction activity of the yeast. In addition, a complicated purification process is necessary when baker""s yeast is used in the production process because it is difficult to separate the yeast from the culture solution after the catalytic reaction.
One embodiment of the invention is a process for producing (6R)-2,2,6-trimethylcyclohexane-1,4-dione. This process includes contacting, in a reactor, 2,6,6-trimethyl-2-cyclohexene-1,4-dione with a yeast selected from the group consisting: of Saccharomyces rouxii (Zygosaccharomyces rouxii), Saccharomyces delbrueckii (Saccharomyces unisporus, Torulaspora delbrueckii), Saccharomyces willianus, Zygosaccharomyces bailii, Candida tropicalis, functional equivalents, subcultures, mutants, and variants thereof, in water, a water-miscible organic solvent, or a mixture of water and the water-miscible organic solvent containing at least one assimilable carbon source; and isolating (6R)-2,2,6-trimethylcyclohexane-1,4-dione produced by the yeast from the reaction medium.
Another embodiment of the invention is a process for producing (6R)-2,2,6-trimethylcyclohexane-1,4-dione. This process includes contacting 2,6,6-trimethyl-2-cyclohexene-1,4-dione with a reaction medium containing yeast, an aqueous solvent, and at least one assimilable carbon source, wherein the yeast is selected from the group of Saccharomyces rouxii (Zygosaccharomyces rouxii), Saccharomyces delbrueckii (Saccharomyces untisporus, Torulaspora delbrueckii), Saccharomyces willianus, Zygosaccharomyces bailii, and Candida tropicalis. The (6R)-2,2,6-trimethylcyclohexane-1,4-dione produced by the yeast is then isolated from the reaction medium.
A further embodiment of the invention is process for producing (6R)-2,2,6-trimethylcyclohexane-1,4-dione. This process includes entrapping yeast cells within a carrier made of a hydrophobic photo-crosslinkable resin having at least two ethylenic unsaturated linkages per molecule, wherein the yeast cells are selected from the group of Saccharomyces rouxii (Zygosaccharomyces rouxii), Saccharomyces delbrueckii (Saccharomyces unisporus, Torulaspora delbrueckii), Saccharomyces willianus, Zygosaccharomyces bailii, and Candida tropicalis. The resin is then irradiated with UV light to form a polymerized yeast carrier. The yeast in the yeast carrier is then preconditioned in a growth medium containing at least one assimilable carbon source, an assimilable nitrogen source, and inorganic salts. 2,6,6-trimethyl-2-cyclohexene-1,4-dione is then added to a reaction medium containing the preconditioned yeast cells from the previous step, an aqueous solvent, and at least one assimilable carbon source. Then, the (6R)-2,2,6-trimethylcyclohexane-1,4-dione produced by the yeast is isolated from the reaction medium.